Gene expression is regulated by a variety of different mechanisms at several different levels. Epigenetic mechanisms regulate gene expression by modifying DNA without changing nucleotide sequences or modifying histones that wrap the DNA molecules and restrict access of DNA binding proteins such as transcription factors. Histone modifications include acetylation, methylation, phosphorylation, ubiquitinylation, and a few others. These modifications are called ‘writing’ and the enzymes responsible for the “writing” are called “writers”. And these histone modifications are reversible, and the enzymes that carry out the reverse mechanisms, such as histone deacetylases and histone demethylases, are called “erasers”.
These epigenetic modifications are typically recognized by so called “readers”, leading to activating or silencing the gene expression depending on specific chromatin context of each writing-reading combination. Bromodomain (BRD)-containing proteins bind to the acetylated histones through the BRDs, which are about 110 amino acids long in length (P. Filippakopoulos, et al., Cell, 2012, 149:214-231). This highly conserved bromodomain, comprised of four antiparallel alpha-helices and two connecting loops, is found in a number of different classes of proteins including histone acetylases, eukaryotic transcription factors and co-regulators, DNA helicases, chromatin-remodeling complexes and others.
Bromodomain and extra terminal domain (BET) proteins are a subfamily of bromodomain-containing proteins that have two bromodomains and one extra-terminal domain (ET). The subfamily is comprised of 4 members including BRD2, BRD3, BRD4, and BRDT (BRD5). BET proteins play an important role in several transcriptional programs, and implicate in aberrant transcriptional events that are responsible for several types of human diseases including inflammation and cancer (A. C. Belkina, et al., Nature Reviews Cancer, 2012, 12 (7):465-477; and R. K. Prinjha, et al., Trends in Pharmacological Sciences, 2012, 33:146-153). Deregulated expression of BRD2, BRD3 and BRD4 is oncogenic in humans. Reciprocal chromosomal translocations between human BRD3 (9q34.2) or BRD4 (19p13.1) genes and the NUT gene (15q14) produce a fused oncoprotein causing an NUT midline carcinoma (NMC), and an aggressive cancer with high mortality (C. A. French, et al., Cancer Research, 2003, 63(2):304-307; and C. A. French, et al., Oncogene, 2008, 27:2237-2242). BRD4 is often up-regulated in melanoma (M. F. Segura, et al., Cancer Research, 2013, 73(20):6264-6276).
Numerous small-molecule inhibitors of BET proteins have been developed which prevent binding of BRD to acetylated histones (S. Muller et al., Med Chem Comm, 2014, 5:288-296). Most of these compounds are acetylated-lysine mimics, and show strong anti-tumor activity against hematological cancers and solid cancers such as mixed lineage leukemia (MLL)-fusion leukemia (Dawson M A et al., Nature, 2011, 478:529-533), multiple myeloma (J. E. Delmore et al., Cell, 2011, 146:904-917, and Aristeidis Chaidos et al., Blood 2014, 123:697-705), glioblastoma (Zhixiang C. et al., Clinical cancer research, 2013, 19:1748-1759), neuroblastoma (J. A. Mertz et al., Cancer Discovery 2013, 3:308-323), prostate cancer (A. Wyce et al., Oncotarget 2013, 4:2419-2429), lung cancer (Shimamura T. et al., Clinical cancer research, 2013, 19:6183-6192), melanoma (M. F. Segura, et al., Cancer Research 2013, 73(20):6264-6276), and autoimmune diseases. Notably, BRD inhibitors inhibited the expression of the oncogene c-myc, which had a critical effect on cell proliferation in many different types of cancers. Therefore, BRD inhibitors or BET protein inhibitors potentially represent a new class of therapeutics to prevent or treat precancerous transformation or a cancer.